The corpora allata (CA) of insects are endocrine glands whose product, juvenile hormone (JH), plays a major role in the control of postembryonic development and reproduction. The CA of Lepidoptera like the tobacco hornworm, Manduca sexta, synthesize at least three homologs of JH, whose titers and ratios vary during development. Because these endocrine glands are directly innervated by axons of cerebral neurosecretory and ordinary neurons, it has been postulated that nervous and/or neuroendocrine regulation of the CA controls fluctuations in the JH titer. For neuroendocrine control, neurohormones could be delivered to the CA either by a paracrine (semi-private; directed) pathway or humorally. Furthermore, in vivo studies have suggested that neuroendocrine control may involve both activating factors (allatotropins) and inhibiting factors (allatohibins). Thus the synthesis of individual homologs could be regulated simultaneously by both allatotropins and allatohibins. The research proposed continues and investigation of the neuroendocrine control of JH synthesis in Manduca. An in vitro approach was developed for these studies to minimize the prospective complexity of this system. This in vitro system utilizes radioimmunoasays for individual JH homologs to monitor the effects of neural factors on JH synthesis by the CA. Two cerebral regulatory factors have been identified with this system, one an allatotropic factor (ATF) specifically stimulating JH III synthesis, and the other an allata-inhibiting factor (AHF) specifically inhibiting JH I synthesis. The JH III ATF is a neuropeptide with an Mr of 40 kd and a pI of 5.5. It is now proposed to purify the JH III ATF, to investigate its mechanism of action and to identify its site of synthesis. The chemical nature of the JH I AHF will be defined and its purification initiated. The in vitro approach will also be used to probe for allatotropins regulating the synthesis of JH I and JH II. This will entail the development of an RIA or JH II and investigation of different brain donors as sources of these factors. The information from these studies should provide a better understanding of the neuroendocrine control of insect postembryonic development and possibly indicate new biological approaches to the control of insect pests. It may also define a system for the investigation of the paracrine regulation of an endocrine gland.